Uric Acid Stimulates Fructokinase and Accelerates Fructose Metabolism in the Development of Fatty Liver

<div><p>Excessive dietary fructose intake may have an important role in the current epidemics of fatty liver, obesity and diabetes as its intake parallels the development of these syndromes and because it can induce features of metabolic syndrome. The effects of fructose to induce fatty liver, hypertriglyceridemia and insulin resistance, however, vary dramatically among individuals. The first step in fructose metabolism is mediated by fructokinase (KHK), which phosphorylates fructose to fructose-1-phosphate; intracellular uric acid is also generated as a consequence of the transient ATP depletion that occurs during this reaction. Here we show in human hepatocytes that uric acid up-regulates KHK expression thus leading to the amplification of the lipogenic effects of fructose. Inhibition of uric acid production markedly blocked fructose-induced triglyceride accumulation in hepatocytes in vitro and in vivo. The mechanism whereby uric acid stimulates KHK expression involves the activation of the transcription factor ChREBP, which, in turn, results in the transcriptional activation of KHK by binding to a specific sequence within its promoter. Since subjects sensitive to fructose often develop phenotypes associated with hyperuricemia, uric acid may be an underlying factor in sensitizing hepatocytes to fructose metabolism during the development of fatty liver.</p> </div

Uric acid sensitizes human hepatocytes to fructose.

<p>(A–B) KHK expression in cells pre-exposed to different amounts of uric acid for 72 hours and further incubated with the same amount of fructose for 24 hours. C) Concentration of TG in liver extracts from cells pre-exposed to different amounts of uric acid for 72 hours and further incubated with the same amount of fructose for 24 hours. D) Adding back uric acid reverts the inhibitory effect of allopurinol on TG accumulation in fructose-exposed HepG2 cells.</p

Allopurinol prevents KHK up-regulation in adult male rats drinking fructose (n = 5 for each group).

<p>(A–B) KHK and AldoB protein expression in liver extracts from control, fructose-fed and fructose with allopurinol-fed rats. C) mRNA expression of <i>khk</i> normalized to <i>β-actin</i> levels in liver extracts from control, fructose-fed and fructose with allopurinol-fed rats. D) Expression of lipogenic proteins FAS, ACC and ACL in liver extracts from control, fructose-fed and fructose with allopurinol-fed rats. E) Expression of the fat oxidation-related protein ECH1 in liver extracts from control, fructose-fed and fructose with allopurinol-fed rats. F) Beta-hydroxybutyrate levels in liver and serum normalized to triglyceride levels. G) Serum fructose levels in control, fructose-fed and fructose with allopurinol-fed rats. H) Beta-hydroxybutyrate levels in control, fructose-fed and fructose with allopurinol-fed rats.</p

Identification in human hepatocytes of ChoRE sites in <i>khk</i> promoter that are activated by fructose and blocked with allopurinol.

<p>A) mRNA expression of <i>khk</i> in cells exposed to fructose in the presence of ChREBP dominant negative (dnMLX). B) KHK activity in cells exposed to fructose in the presence of ChREBP dominant negative (dnMLX). C) ChIP analysis and <i>khk</i> promoter occupancy in distal and proximal ChoRE sites by ChREBP in cells exposed to fructose in the presence of ChREBP dominant negative (dnMLX) or allopurinol. D) Luciferase expression in human hepatocytes transfected with pGL3-<i>khk</i>proximal ChoRE and exposed to fructose in the presence of ChREBP dominant negative (dnMLX) E) Luciferase expression in human hepatocytes transfected with pGL3-<i>khk</i>distal ChoRE and exposed to fructose in the presence of ChREBP dominant negative (dnMLX) or allopurinol.</p

Allopurinol prevents fructose-induced CHREBP acetylation and nuclear translocation in human hepatocytes.

<p>A) KHK mRNA expression in cells exposed to fructose (5 mmol/L) for different time points. B) Analysis of acetylation state in immunoprecipitated ChREBP in cells exposed to glucose, fructose or mannitiol in the presence or absence of allopurinol. (C–E) ChREBP and KHK expression in nuclear and cytoplasmic extracts of cells control and incubated with glucose (25 mM) and fructose (5 mM) in the presence or absence of allopurinol.</p